Immunological tolerance is a desirable goal in clinical transplantation as it permits graft acceptance without the need for continuous immunosuppression. A promising experimental tolerance strategy that is currently being tested in humans is the induction of mixed hematopoeitic chimerism. The major obstacle impeding the clinical success of mixed hematopoetic chimerism has been the need for toxic conditioning (myeloablation, lymphodepletion, and/or immunosuppression) of the recipient to ensure donor stem cell engraftment and stable chimerism. In the absence of such conditioning, chimerism and tolerance are rarely achieved. Therefore, a better understanding of what prevents mixed hematopoeitic chimerism is needed to develop safer clinical strategies. The goal of this exploratory grant proposal is to identify the innate immune allorecognition mechanisms that prevent successful chimerism. To do so, we propose to study an invertebrate model organism, Hydractinia symbiolongicarpus, which lacks an adaptive immune system yet exhibits a highly specific innate allorecognition response. The Hydractinia allorecognition system is responsible for preventing chimerism between histo-incompatible animals. The specific aims are: 1. To identify and characterize Hydractinia allorecognition loci. 2. To characterize the stability of allogeneic Hydractinia chimeras. Uncovering innate immune recognition systems in an invertebrate model organism such as Hydractinia is potentially of great relevance to vertebrate immunology as recently illustrated by the discovery of Toll receptors in Drosophila. The identification of Toll-like receptors (TLR) in mammals provided a compelling explanation of how the immune system recognizes and reacts to microbial non-self. Understanding how the innate immune system recognizes allogeneic cells (non-microbial non-self) may be of comparable important biomedical relevance to the field of transplantation. Lay Summary: The success of transplantation in humans is hindered by rejection of the transplanted organ. Understanding the earliest events that trigger rejection is essential for developing effective therapies. Here we propose to use a simple marine organism endowed with the ability to reject unrelated individuals to uncover the most basic events that initiate the rejection process. [unreadable] [unreadable] [unreadable] [unreadable]